Abstract 1639: Strong synergy with APR-246 and DNA-damaging drugs in primary ovarian cancer cells

Abstract

Abstract Background: Mutations in the TP53 gene occur in at least 60% of ovarian tumors and are associated with chemoresistance and poor prognosis. APR-246 (PRIMA-1MET) is the first mutant p53-reactivating compound in clinical development and has been tested as monotherapy in hematological malignancies and prostate cancer with promising results (Lehmann et al. J Clin Oncol 30, 2012). The aim of this study was to investigate the anticancer effects of APR-246 in combination with conventional chemotherapy in cancer cells isolated from ascites fluid from ovarian cancer patients. Methods: Ascites cells were purified by Ficoll and viably frozen, and the quality and purity were confirmed by May Grünwald/Giemsa staining. For some samples, immunocytochemical stainings with anti-Ber-EP4 and anti-calretinin antibodies were used to distinguish between mesothelial and cancer cells. Cell viability was assessed with FMCA assay and Combination Index (CI) calculated using Additive model. CI < 0.8 indicates synergy and CI < 0.5 strong synergy. TP53 gene status was determined by Sanger sequencing and single strand conformation analysis, and p53 protein expression by Western blotting. Results: Eight of ten samples tested were from patients with recurrent ovarian cancer previously treated with platinum drugs. Cancer cells from seven patients possessed TP53 core domain missense mutations L111Q, C135Y, P151H, Y163H, C238F, P278R and R280K, respectively; two had nonsense mutations E346* and E204*, and one was wild type. All the missense mutations have been predicted to severely affect p53 tumor suppressor function. Missense mutant p53 proteins were expressed at high levels while no p53 expression was detected in cells with wild type or nonsense mutant p53. Synergistic or strong synergistic effects with APR-246 and cisplatin were observed in all ten samples tested. Synergy was also observed with the platinum analogue carboplatin and the anthracycline doxorubicin. The IC50 values for cisplatin ranged from 3 to 40 μM and for APR-246 from 5 to 37 μM. We also tested the ability of APR-246 to sensitize the primary ovarian cancer cells carrying Y163H mutant p53, to cisplatin; the IC50-value of cisplatin decreased from 10 to 2.6 μM in the presence of 6 μM APR-246. Conclusions: We observed striking synergy with APR-246 and platinum drugs or doxorubicin. These results are consistent with our previous results in ovarian cancer cell lines showing synergy not only in p53 mutant but also in p53 null cancer cell lines. In these cells, the synergy may be related to the fact that APR-246 decreases intracellular glutathione level. Our results provide a strong rationale for the ongoing clinical study with APR-246 in combination with carboplatin and doxorubicin in patients with recurrent ovarian cancer and suggest that combination treatment with APR-246 and DNA-damaging drugs could allow significantly improved treatment for ovarian cancer carrying mutant p53. Citation Format: Åsa Fransson, Daria Glaessgen, Jessica Alfredsson, Klas G. Wiman, Svetlana Bajalica Lagercrantz, Nina Mohell. Strong synergy with APR-246 and DNA-damaging drugs in primary ovarian cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1639. doi:10.1158/1538-7445.AM2015-1639